Development of specific probes for detection of ropiness causing lactic acid bacteria in cider

IDOIA IBARBURU; MARÍA LAURA WERNING,; GAIZKA GARAI-IBABE; ANA IRASTORZA; MARÍA TERESA DUEÑAS; JESÚS NAVAS; PALOMA LÓPEZ

Rennes

Simposio; EUROPEAN SYMPOSIUM ON APPLE PROCESSING; 2005

Abstract Exopolysaccharide (EPS) production by some lactic acid bacteria have a deletereous effect on texture of cider, wine and beer. This spoilage of drinks is called oiliness or ropiness. The cider becomes heavy, it ropes when poured and flows without any noise. Therefore, a method for detection of EPS-producing bacteria could be useful for prevention of ropy spoilage in drinks. We have previously isolated ropy lactic acid bacteria from spoiled ciders, belonging to the genus Lactobacillus (1), Pediococcus (2) and Oenococcus. Structural characterization of the exopolysaccharide produced by Pediococcus damnosus 2.6 revealed that this strain synthesises a 2-substituted-(1-3)- β-D-glucan (3). In this work, the glycosyltransferase (gtf ) gene from P. damnosus 2.6 has been cloned and its DNA sequence determined. The gtf gene product is homologous to the members of family 2 glycosyltransferases and its catalytic domain is flanked by transmembrane regions. Primers were designed to amplify either the N-terminal transmembrane or catalytic domains encoding regions. PCR analysis using these primers allowed to demonstrate that the Lactobacillus sp. G77, which produces the same EPS than P. damnosus 2.6 (4), also carries the gft gene. Southern analysis revealed that in both bacteria the gene is extrachromosomal. However, a large (35 kb) or small (5,5 kb) plasmid carries the gene in P. damnosus or Lactobacillus. A protocol for direct PCR analysis of colonies has been developed. Its usage has allowed identification of new Pediococcus, Lactobacillus and Oenococcus ropy strains isolated from ropy cider, which have the gtf gene. Finally, a method for detection of ropy bacteria in cider has been standardised.